Aerobic organisms are constantly exposed to endogenous and exogenous reactive oxygen species (ROS). Endogenous ROS include hydroxyl radicals that are produced predominantly as byproducts of normal cellular metabolism and through leakage of electrons during the mitochondrial electron transport chain. The interaction of ROS with the cellular genome often results in oxidatively induced DNA damage that includes; DNA base and sugar modifications, DNA-protein cross-links, abasic sites, single-strand breaks (SSB) and double-strand breaks (DSB). One of the most common oxidatively induced DNA base modifications is 8-OH-G which mispairs with adenine during DNA replication causing GC-TA transversion mutations. To maintain proper genetic integrity and to minimize cancer risk, organisms ranging from E. coli to humans have evolved elaborate mechanisms for repairing 8-OH-G. The E. coli MutT, MutM, and MutY pathways provide efficient repair of 8-OH-G. MutT and its human homologue hMTH is an 8-OH-GTpase that cleanses the nucleotide pool of 8-OH-GTP thus preventing its incorporation into the DNA during DNA replication. MutY and its human homolog hMYH is a monofunctional DNA glycosylase that removes adenine opposite 8-OH-G. MutM (Fpg) is a DNA glycosylase/AP lyase that specifically removes 8-OH-G opposite cytosine. In humans, 8-OH-G is repaired predominantly by hOGG1, a DNA glycosylase/AP lyase that is specific for 8-OH-G and 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG) opposite cytosine via the base excision repair (BER) pathway. There is a growing body of literature associating oxidative DNA damage with cancer. This includes work showing that the level of hydroxyl radical-induced DNA damage is increased in DNA of invasive ductal breast carcinoma tissues relative to normal breast tissue, implicating 8-OH-G repair in breast cancer development. Olinski and coworkers have also shown that free-radical-induced DNA damage is elevated in chromatin of various surgically removed cancerous tissues from the colon, stomach, ovary, brain and lung relative to the surrounding non-cancerous tissues. Peripheral blood lymphocytes of women at high risk of developing breast cancer were shown to be deficient in the repair of radiation-induced DNA damage. Furthermore, mitochondrial extracts of breast cancer cell lines, MCF-7 and MDA-MB-468 are defective in the repair of 8-OH-G. To date the causes of the defective repair in these cancer cell lines remains unresolved. We analyzed the ability of HCC1937 and MCF-7 breast cancer cells to repair this lesion relative to the nonmalignant human mammary epithelial cell line, AG11134. We show that HCC1937 cells have diminished nuclear repair of 8-OH-G relative to AG11134 and have no detectable hOGG1. Nuclear DNA of HCC1937 and MCF-7 cells accumulated higher levels of 8?-hydroxyl-2?-deoxyguanosine (8-OH-d-G) after H2O2 treatment followed by a repair period compared to AG11134 cells. The deficient repair is not associated with a genetic change in the hOGG1 gene since the coding and the promoter regions of this gene did not harbor any mutations. Repair of 8-OH-G in HCC1937 cells was significantly stimulated by purified hOGG1. Furthermore, expression of hOGG1 by transfection of the hOGG1 gene in HCC1937 cells resulted in increased repair of 8-OH-G. HCC1937 cells had significant upregulation of SOD1 and SOD2 and exhibited reduced clonogenic survival after oxidative stress. This study provides evidence for inefficient in vivo and in vitro repair of 8-OH-G in HCC1937 human breast cancer cells and directly implicates hOGG1 in this defect.